One of the major objectives is to purify the enzyme involved in the N-glycosylation of proteins, oligosaccharide transferase. The specificity of the purified enzyme toward 1) saccharide-lipid substrates, ranging in size from Glc3-Mang-GlcNAc2-PP-Dolichol to GlcNAc-PP-Dolichol, and 2) Asn-X-Ser/Thr substrates in which the X group is systematically varied, will be studied. Potential reversible inhibitors of the transferase, consisting of Asn-Leu-Thr modified in the amide side of Asn will be prepared. These modified peptides or Asn-Leu-Thr will be converted to potential irreversible inhibitors by attachment of active site reactive probes. These inhibitors of oligosaccharide transferase will be utilized in vitro and in vivo studies that should cast light on 1) regulation of the pathway of oligosaccharide-lipid synthesis and 2) the possible role of glycosylation in regulating translation. The second major objective is to investigate the function of another class of polyisoprenoids, the retinoids. Because it is clear that retinol is essential for spermatogenesis, we will study two testes-derived cell types, the Sertoli and Leydig cells. The effect of added retinol or retinoic acid on macromolecular synthesis in these cells will be studied. Particular emphasis will be given to the possible effect of retinol on glycoconjugate synthesis. The possibility that mannosylphosphorylretinol serves as a mannosyl donor in glycoprotein synthesis will be examined both in vivo and in vitro. The oligosaccharide chain of the mannose labeled proteins will be characterized and the steps in assembly of these chains will be studied. Since studies with several cell types suggest that retinoic acid acts in a manner analogous to steroid hormones, the possibility that retinoic acid stimulates production of specific secretory proteins will be studied and the molecular basis of the stimulation will be investigated.